**3.1 Anti-biofilm activity of meta-bromo-thiolactone (mBTL) via QS inhibition**

In the study of O'Loughlin et al. [13], synthetic molecules were analyzed to prove their inhibitor effects on the two *P. aeruginosa* quorum-sensing receptors, LasR and RhlR. It was found that the most effective compound, is the meta-bromo-thiolactone (mBTL). It was also confirmed that both LasR and RhlR are partially inhibited by mBTL *in vivo* and *in vitro*; however, RhlR, not LasR, is the relevant *in vivo* target. Therefore, this work, that explores interference with QS, demonstrates that mBTL, an analogue of the native self-inducers of *P. aeruginosa*, suppresses the expression of genes encoding the virulence factor pyocyanin, on the one hand, and prevents biofilm formation on the other hand, which protects *C. elegans* and human lung epithelial cells from attack by *P. aeruginosa.* Taken together, these data about mBTL provide a strong argument for the efficacy of QS modulators for attenuation of QS-controlled phenotypes in pathogenic bacteria, such as biofilm formation (**Table 1**).

## **3.2 Anti-biofilm effect of malondialdehyde (MDA) via cell membrane injury**

Malondialdehyde (MDA), one of the most representative reactive carbonyl species (RCSs) produced by lipid oxidation in bacteria [19] and in food [14], has received extensive attention recently. However, the inhibitory effect of MDA on microorganisms has received little attention. The study of Zhang et al. [10] proved the antibacterial effects of MDA on *S. xylosus* and *Lactiplantibacillus plantarum* with the MICs of 90 and 180 μg/ml, respectively. In addition, the antibacterial mechanisms of MDA on these two bacteria were associated with LDH activity changes as the LDH release is indicator of cell wall injury, accompanied with Ca2+ and Mg2+ leakage. Overall, the emission of Ca2+ and Mg2+ demonstrated that MDA enhanced the permeability of *S. xylosus* and *L. plantarum* cell membrane and further affected bacterial metabolism. In addition, MDA treatment induces cell membrane depolarization, indicating severe membrane damage with important impact on cell development and differentiation. This result has been confirmed by combination of CLSM and FEGSEM observations which have affirmed that MDA disrupts the cell membrane of *S. xylosus* and *L. plantarum.* It was also shown that MDA treatment significantly reduced the ATP concentration in *S. xylosus* and *L. plantarum,* suggesting that MDA may inhibit their growth by affecting the metabolic functions or cell membrane permeability of bacteria. Moreover, FT-IR studies showed that MDA might affect the molecular composition of *S. xylosus* and *L. plantarum* cells. These changes indicated the negative influence of MDA on cell membrane and cellular homeostasis [14].
